Neonatal Sepsis Episodes and Retinopathy of Prematurity in Very Preterm Infants

This cohort study investigates the association of neonatal sepsis episodes with retinopathy of prematurity, including treatment-warranted cases, among very preterm infants in Germany and Norway.


Introduction
2][3] Timely diagnosis and treatment are essential to prevent irreversible visual impairment or blindness.Low gestational age (GA), low birth weight, poor postnatal growth, and exposure to hyperoxia and hypoxia are well-known risk factors. 1,3,4With improved survival of very preterm infants and despite an ongoing search for optimized strategies to control oxygen saturation and supply, [5][6][7] rates of ROP have not declined. 2,3,8e incidence of treatment-warranted ROP in overall screening cohorts of very preterm infants in high-income countries ranges from 2% to 6%. 3,9 In selected high-risk cohorts, an incidence of 4% to 15%, sometimes as high as 30%, is reported. 8,9Laser photocoagulation and intravitreal antivascular endothelial growth factor (VEGF) injections are the most prevalent treatment modalities. 1,3,10,11Of note, even infants with timely treatment or ROP stages below treatment thresholds are at increased risk of permanent visual impairment. 1,3Therefore, a better understanding of actionable risk factors for prevention and early diagnosis remains paramount.
0][21][22][23] Small prospective studies support these findings. 24,25Whether early-onset sepsis (EOS) and (recurrent) late-onset sepsis (LOS) are associated with different changes in ROP risk remains unclear. 15,26,27In this study, we used data from the German Neonatal Network (GNN) and Norwegian Neonatal Network (NNN) to test whether very preterm infants who survived 1 or more episode of culture-proven sepsis were at increased risk of developing ROP or treatment-warranted ROP.

Study Population
The GNN is a population-based observational multicenter study involving 68 level III neonatal intensive care units (NICUs) in Germany investigating risks and complications of infants with very low birth weight born at a gestation of 22 weeks and 0 days to 36 weeks and 6 days.This study includes infants born at 22 weeks and 0 days to 28 weeks and 6 days enrolled between January 1, 2009, and December 31, 2022.Written informed consent by parents or guardians was mandatory for enrollment.Predefined clinical data were recorded on case report forms, coded, and stored centrally at the coordinating center in Lübeck, Germany.All infants born at less than 29 weeks admitted to participating centers were eligible.Exclusion criteria were lethal malformation and lack of written consent.All study parts of the GNN were approved by the ethics committee of the University of Lübeck and local ethics committees of all participating centers.GNN review and satisfaction of informed consent extend to this study.Data quality was ensured by annual on-site monitoring.The NNN is a national population-based registry comprising all 21 NICUs across 4 health trust regions.
Reporting to the registry is compulsory without consent within current national legislation and includes daily registrations of all investigations, treatments, and diagnoses of all infants admitted to 1 or several Norwegian NICUs until final discharge. 28,29The NNN cohort in this study includes infants born at a gestation of 22 weeks and 0 days to 28 weeks and 6 days between January 1, 2009, and

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Neonatal Sepsis Episodes and Retinopathy of Prematurity in Very Preterm Infants and health research ethics.NNN review and exemption for informed consent extend to this study.
Our study followed the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) reporting guideline for cohort studies.

Ophthalmological Screening and ROP Definition
In Germany and Norway, standardized ROP screening during the study period included all preterm infants born at less than 32 weeks' gestation (<31 weeks in Germany from 2020), with birth weight less than 1500 g independent of the need of supplemental oxygen, or both characteristics.Moreover, screening is recommended in Germany for infants born at a gestation of 31 weeks and 0 days to 36 weeks and 6 days with a requirement of supplemental oxygen for more than 5 days or the presence of severe comorbidities, such as necrotizing enterocolitis (NEC), bronchopulmonary dysplasia, postnatal sepsis, or severe anemia requiring red blood cell (RBC) transfusion. 30In GNN and NNN centers, screening started at ages 5 to 6 weeks and not before age 31 weeks.Examinations were performed weekly or biweekly depending on individual findings and were continued until physiologic vascularization reached zone III, preexisting ROP showed continuous regression, or both conditions were met.ROP was defined according to the International Classification of Retinopathy of Prematurity consensus statement. 31Severity was designated by zone, stage, and circumferential extent assessed by sectors and the absence or presence of plus disease. 31Treatment-warranted ROP referred to aggressive ROP, any stage ROP in zone I with plus disease, ROP 3 in zone I, or ROP 3+ in zone II according to national guidelines that required laser therapy, cryotherapy, or anti-VEGF treatment. 30,32

Sepsis Definition
Neonatal sepsis was defined as blood culture-proven sepsis identified by clinical sepsis criteria and detection of a causative pathogen in 1 or more blood cultures according to criteria of the German National Nosocomial Infection Surveillance System in Preterm Infants (NEO-KISS). 33EOS was defined as sepsis occurring within the first 72 hours of life, while late-onset sepsis (LOS) was defined as sepsis episode after 72 hours of life. 34,35The analysis of 1, 2, or 3 episodes included EOS or LOS episodes.Neither the GNN nor the NNN dataset provided data on the respective causative agent.

Definitions of Clinical Parameters
GA was calculated from the best obstetric estimate according to early prenatal ultrasonography and obstetric examination.Small for GA was defined as a birth weight less than the 10th percentile according to GA; inborn was defined as birth in a network center.Clinical chorioamnionitis, with data available only from GNN, was documented by the attending obstetrician as cause of preterm birth based on maternal fever plus fetal tachycardia, a white blood cell count greater than 15 000/μL, or a foul-smelling discharge.Days on supplemental oxygen and days on mechanical ventilation or continuous positive airway pressure (CPAP) referred to the number of complete days of exposure.
Bronchopulmonary dysplasia was defined as requirement of oxygen therapy for at least 28 days and classified as mild, moderate, or severe according to the need for supplemental oxygen at 36 weeks' postmenstrual age. 36Intraventricular hemorrhage was diagnosed according to Papile classification. 37stic periventricular leukomalacia was defined as periventricular, cystic white matter lesions.Both entities were diagnosed using cranial ultrasonography.NEC requiring surgery corresponded to clinical NEC classified as Bell stage II or III with the need for laparotomy 38 and macroscopic diagnosis of NEC.

Statistical Analysis
Study groups were compared using univariable analyses, including the Mann-Whitney U test for continuous and 2-sided χ 2 test for categorical variables.The primary analysis used stepwise multivariable logistic regression to adjust for confounders, accounting for the multihit sequence of  1).EOS was not associated with ROP risk.On the contrary, 1 sepsis episode occurring as LOS and 2 to 3 episodes of sepsis as EOS plus LOS or recurrent LOS were associated with increased ROP risk (Table 1).
Comparisons further suggested that infants with ROP had a lower GA and birth weight and were more likely to be small for GA.Moreover, this group spent more days on mechanical ventilation or CPAP and supplemental oxygen, had a higher maximum fraction of inspired oxygen in the first 12 hours, needed more days until full enteral feeds, and was more likely to have a history of inotropes in the first 24 hours and of intraventricular hemorrhage, periventricular leukomalacia, and NEC requiring surgery.Antenatal steroids and less invasive surfactant administration were associated with reduced ROP risk (Table 1).
Using a stepwise logistic regression approach, we adjusted for known and probable risk factors of sepsis with odds of treatment-warranted ROP vs 0 episodes (Table 3; eFigure 3 in Supplement 1).
Given the interrelated nature of variables included in this modeling, we performed propensity score matching as an alternative statistical approach.In 2122 matched pairs of infants without sepsis and infants with 1 sepsis episode, sepsis was associated with ROP (aOR, 1.76 [95% CI, 1.54-2.02];P = .007)sepsis episodes, recurrent sepsis was associated with increased odds of ROP vs 0 or 1 episode (Table 3).There were increasing rates of treatment-warranted ROP with each sepsis episode  Treatment-warranted ROP

B
The total number of infants in the respective gestational age (GA) strata and sorted by number of sepsis episodes was as follows: For infants with a GA of 22 weeks and 0 days to 24 weeks and 6 days, there were 1831 infants with 0, 588 infants with 1, 155 infants with 2, and 50 infants with 3 episodes of sepsis.For infants with a GA of 25 weeks and 0 days to 26 weeks and 6 days, there were 3776 infants with 0, 702 infants with 1, 112 infants with 2, and 28 infants with 3 episodes of sepsis.For infants with a GA of 27 weeks and 0 days to 28 weeks and 6 days, there were 5023 infants with 0, 468 infants with 1, 50 infants with 2, and 11 infants with 3 episodes of sepsis.
( infants (2122 matched pairs of study infants without sepsis and infants with 1 sepsis episode) for the analysis of 1 sepsis episode and 800 infants (400 matched pairs of infants without sepsis or 1 sepsis episode and infants with 2-3 episodes) for the analyses of 2 or 3 sepsis episodes.Both pairs showed full matching defined as no differences between demographic and clinical characteristics.Propensity score matching was performed using gestational age, small for gestational age, sex, clinical chorioamnionitis, and presence of central venous line as variables, all associated with the individual's risk of sepsis.Subsequent conditional regression analysis adjusted for variables included in the stepwise logistic regression modeling.
c The reference group for all aORs is infants with 0 sepsis episodes except for analyses of 2 or 3 sepsis episodes in propensity score-matched infants, for which the reference group was infants with 0 or 1 sepsis episode.a Variables not associated with treatment-warranted ROP in univariable analysis were female sex, intraventricular hemorrhage grade 3 to 4, and cystic periventricular leukomalacia.
b The reference group was infants with 0 sepsis episodes.
c Outcomes were per 1-week increase in GA.

Discussion
In this cohort study, we found an association of neonatal sepsis and recurrent episodes with an increased risk of ROP and treatment-warranted ROP in a large dataset from Germany.26][27]40 To the best of our knowledge, our study is the first to describe a dose-response association between the number of sepsis episodes and increases in the odds of ROP and treatment-warranted ROP.Notably, in the GNN dataset culture-proven sepsis was associated with a greater increase in risk of ROP and treatment-warranted disease than were GA, birth weight, and days on supplemental oxygen, although these have been widely acknowledged as major risk factors. 1,3,4,8,25In the smaller cohort from Norway, there was no association between neonatal sepsis and treatment-warranted ROP after adjusting for confounders.Instead the inflammatory condition of NEC was associated with the greatest increase in risk of treatmentwarranted ROP in the NNN cohort.
This observational study found associations and results cannot be used to conclude causation. 4114][15][16][17]39 Underlying mechanisms remain to be elucidated. 39,443][47][48][49][50][51] Inflammation-induced reactive oxygen species and oxidative stress, as well as inflammation-induced VEGF expression, microglia activation, and retinal ganglion cell death have been discussed as causative mechanisms. 39,44,47,48In rat models, microglia-derived interleukin (IL)-1β was found to induce retinal ganglion cell death and breakdown of the blood-retina barrier, 47,52 while application of IL-1 receptor antagonists attenuated vasoobliteration. 53In interaction with growth factors, a number of other leukocyte and microglia-derived cytokines and chemokines have been implicated in the disturbance of retinal vasculature. 54,55Ongoing basic research may help to identify therapeutics modulating inflammation-driven retinal damage. 54,55Notably, animal models showed that intravitreal administration of noncoding RNAs and gut microbiota modulation were potentially effective in regulating the course of ROP. 55The latter approach is in line with increasing evidence for a gut-retina axis and increasing interest in understanding the potential associations of infections, use of antibiotics, and gut dysbiosis with the development of retinopathy. 56Ultimately, hemodynamic and respiratory instability, frequently accompanying sepsis and NEC, may impair tissue perfusion and oxygen saturation and cause retinal ischemia at a vulnerable stage of development. 39,57In summary, neonatal sepsis and systemic inflammation observed in NEC may be closely intertwined in a multihit sequence of ROP. 19,39,44In this context, it may be worthwhile to discriminate whether multiple sepsis episodes are surrogates for true new infections or instead for recurrent episodes of the same origin or correlative of sustained inflammation with an undulating course.
While some studies have indicated that LOS but not EOS was associated with increased risk of ROP, 15,26,58 others found an association for EOS and yet others for infections early and later in life. 27,59In a meta-analysis, 16 EOS and LOS were associated with severe ROP.However, only 3 of 34 studies discriminated between EOS and LOS. 16In our study, EOS alone was not associated with ROP or treatment-warranted ROP, while repetitive LOS episodes or EOS followed by LOS were associated.
Whether prenatal exposure to inflammation, namely chorioamnionitis, and subsequent fetal inflammation are associated with increased risk of later ROP has not been conclusively answered. 4,46,49,60,61An inflammation-induced prenatal sensitization of the immature retina to secondary injurious hits has been discussed. 1Two meta-analyses 22,62 evaluating chorioamnionitis and ROP development found no association with ROP or treatment-warranted ROP when adjusting for confounders.A 2024 meta-analysis 63 described an association between histological chorioamnionitis and the development of severe ROP.Notably, these findings were confounded by GA. 63 On the contrary, a 2023 study 64 in 182 placental tissues of infants born at less than 32 weeks and with a birth weight less than 1500 g that included comprehensive histopathological examinations and plasma protein analyses found that acute placental inflammation was associated with reduced risk of ROP.In addition, a 2024 nation-based cohort study 65 from Japan comprising 38 013 preterm infants born at 1500 g or less or less than 32 weeks of gestation found a reduced risk of severe ROP in infants with severe histological chorioamnionitis.In keeping with these findings, a history of clinical chorioamnionitis was associated with decreased odds of ROP and treatmentwarranted ROP in our study.Data on histological exams of placentas were not available.
The prevalence of any ROP in GNN and NNN study cohorts was 49.8% and 33.6%, respectively.7][68][69] Whether diverging ranges reflect differences in active care and survival of the most premature infants, mirror varying screening routines, or indicate risk factors not sufficiently addressed in some centers remains to be elucidated.
Regardless of the case and against the background of an ever-decreasing GA of today's NICU population, these numbers highlight the need to continue deciphering underlying mechanisms of ROP and disease progression and to improve prevention.1][72] So far, neonatal sepsis has not been included in any of these scores.

Limitations
This study has some limitations.These are partly related to the observational design that does not allow for any causal conclusion and may result in residual confounding.In addition to unknown differences in baseline characteristics, differences in clinical care or poor overall clinical course may affect rates of ROP and treatment-warranted ROP.Addressing these concerns, we controlled for several known and probable prenatal, perinatal, and infant risk factors, including NEC requiring surgery and RBC transfusions, which were also associated with ROP and treatment-warranted ROP.
4][75][76] On the contrary, our analyses confirmed previous findings of protective associations of antenatal steroids with ROP development. 77Moreover, female sex was associated with decreased odds of treatmentwarranted ROP.This observation may be in line with the frequently observed phenomenon that more male than female infants are treated for ROP. 8,78It is imperative to mention that there may be potential confounders that were missed in applied regression models.The GNN dataset registered largely stable rates of ROP between 2009 and 2022.However, the evolution of neonatal care of very preterm infants in this 14-year period could have biased the results.Additionally, our study was limited by the relatively small sample size of infants with treatment-warranted ROP despite the use of 2 large-scale datasets spanning a 14-year and 10-year observation period.Findings in both datasets were in the same direction.However, large and well-powered prospective studies are needed to confirm our findings.

Figure .
Figure.Rates of Retinopathy of Prematurity (ROP) in the German Neonatal Network

Table 2 .
Characteristics of Infants in the Norwegian Neonatal Network

Table 3 .
Association of Neonatal Sepsis With ROP in the German Neonatal Network a Stepwise multivariable regression modeling included 6424 infants without ROP and 6370 infants diagnosed with any ROP.See the eMethods in Supplement 1 for descriptions of each model.bConditional regression analysis in propensity score-matched infants included: 4244

Table 4 .
Association of Neonatal Morbidities With Treatment-Warranted ROP in the Norwegian Neonatal Network

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Sepsis Episodes and Retinopathy of Prematurity in Very Preterm Infants JAMA Network Open.2024;7(7):e2423933.doi:10.1001/jamanetworkopen.2024.23933(Reprinted) July 25, 2024 8/15 Downloaded from jamanetwork.comby guest on 07/27/2024 Neonatal Sepsis Episodes and Retinopathy of Prematurity in Very Preterm Infants honoraria from Pfizer, Merck Sharp and Dohme, Sanofi, and AstraZeneca outside the submitted work; holding a patent with Chiesi for less invasive surfactant therapy; and having membership on a board of the German Research Foundation.Dr Fortmann reported receiving speaker honoraria from Chiesi and Pfizer.Dr Nentwich reported receiving speaker honoraria from Novartis.Dr Stahl reported receiving grants from Novartis, Bayer, and Roche and personal fees from Novartis, Bayer, Alcon, Apellis, and Roche outside the submitted work.Dr Göpel reported receiving personal fees from Chiesi and Abbott outside the submitted work.No other disclosures were reported.77.Higgins RD, Mendelsohn AL, DeFeo MJ, Ucsel R, Hendricks-Munoz KD.Antenatal dexamethasone and decreased severity of retinopathy of prematurity.Arch Ophthalmol.1998;116(5):601-605. doi:10.1001/archopht.116.5.601 78.Hoyek S, Peacker BL, Acaba-Berrocal LA, et al.The male to female ratio in treatment-warranted retinopathy of prematurity: a systematic review and meta-analysis.JAMA Ophthalmol.2022;140(11):1110-1120. doi:10.1001/jamaophthalmol.2022.3988Study Flowchart of Infants Enrolled in the German Neonatal Network eFigure 2. Study Flowchart of Infants Enrolled in the Norwegian Neonatal Network eFigure 3. Variables Independently Associated With ROP in GNN in Stepwise Multivariable Regression Analyses eTable 1. Multivariable Logistic Regression Modeling Testing for Association Between Neonatal Sepsis and Development of Any ROP in Very Preterm Infants in the GNN eTable 2. Multivariable Logistic Regression Modeling Testing for Association Between Neonatal Sepsis and Development of Treatment-Warranted ROP in the GNN eMethods.Description of Models Used in Analysis of Association With ROP and Treatment-Warranted ROP in GNN